by
k66 » Fri Aug 24, 2007 7:18 am
"a C18 is not a C18 is not a C18"
In the early days of HPLC, octadecylsilane (most often referred to as an ODS or C18 phase) was one of the first bonded stationary phases that became available for the new technique called "reversed-phase chromatography." It became the standard phase for reversed-phase chromatography and quickly was adopted by most practitioners. Because the pharmaceutical industry was an early adopter of HPLC and regulatory bodies did not want to bless a particular manufacturer's brand of column, the FDA and USP developed a classification system that gave a generic designation for each new method that was submitted under a new drug application. For HPLC columns, an "L" designation was given and because C18 was used in a majority of the submittals, it became "L1." As additional phases were added, they were given their own "L" number (for example, L7 C8, L10 CN, L11 Phenyl, and so forth).
Unfortunately, this designation system proved to be unreliable because each commercial C18 column was synthesized differently using silica gel as the base material and the resulting reversed-phase columns had different properties. For example, some manufacturers used octa-decylmonochlorosilane and a low surface area silica gel (Figure 1) while others used the same silane but bonded it to a higher surface area silica gel. These two C18 columns would behave differently with the latter having more C18 phase than the former. A lower coverage bonded phase sometimes had unreacted silanols that caused mixed retention mechanisms. Some manufacturers use di- and trichlorosilanes and polymerized the bonded phase to form a thicker layer with different diffusional properties. To cut down on unreacted silanols that remained after the bonding process, some manufacturers endcap these silanols with a small silane (for example, trimethylmonochlorosilane). Silanols are sometimes responsible for tailing of basic compounds at intermediate pH values. Some manufacturers even went further and double-endcapped with a second small silane to provide a more inert surface. Some manufacturers used polymeric base materials and then bonded C18 moieties to their surface resulting in a totally different C18 packing material, yet it was still considered an "L1." Others use organoalkoxysilanes that display a different degree of reactivity and can produce a slightly different C18 bonded phase than would an organochlorosilane reagent.
So, the statement "a C18 is not a C18 is not a C18" holds true even today. All C18 phases are not alike and the user should make sure that once a method is developed, he or she sticks with the same column part number to ensure rugged and robust methods. In recent times, Dolan and Snyder (1,2) have studied a large number of C18 phases in an attempt to determine which phases have similar retention and selectivity features. By characterizing the phases based upon probes with as many as five retention parameters, they have classified phases with a numerical value so that one might be able to choose the closest phase in performance characteristics in case the original phase is not available from a particular manufacturer. Their approach also allows one to find a C18 phase totally unlike their existing phase that can aid in column selection during method development.
I have never used an ACE column except for Cyano ones (and they are pretty good actually).